Genomics and morphometrics reveal the adaptive evolution of pikas.

Pikas (Lagomorpha: Ochotonidae) are small mouse-like lagomorphs. To investigate their adaptation to different ecological environments during their dispersal from the Qinghai-Xizang (Tibet) Plateau (QTP), we collected 226 pikas and measured 20 morphological characteristics and recorded habitat information. We also sequenced the genome of 81 specimens, representing 27 putative pika species. The genome-wide tree based on 4 090 coding genes identified five subgenera, i.e., Alienauroa, Conothoa, Lagotona, Ochotona, and Pika, consistent with morphometric data. Morphologically, Alienauroa and Ochotona had similar traits, including smaller size and earlier divergence time compared to other pikas. Consistently, the habitats of Alienauroa and Ochotona differed from those of the remaining subgenera. Phylogenetic signal analysis detected 83 genes significantly related to morphological characteristics, including several visual and hearing-related genes. Analysis of shared amino acid substitutions and positively selected genes (PSGs) in Alienauroa and Ochotona identified two genes, i.e., mitochondrial function-related TSFM (p.Q155E) and low-light visual sensitivity-related PROM1 (p.H419Y). Functional experiments demonstrated that TSFM-155E significantly enhanced mitochondrial function compared to TSFM-155Q in other pikas, and PROM1-419Y decreased the modeling of dynamic intracellular chloride efflux upon calcium uptake. Alienauroa and Ochotona individuals mostly inhabit different environments (e.g., subtropical forests) than other pikas, suggesting that a shift from the larger ancestral type and changes in sensory acuity and energy enhancement may have been required in their new environments. This study increases our understanding of the evolutionary history of pikas.

A theoretical design method for the nozzle pitch of air-flotation system to reduce large liquid crystal display glass substrate's deformation.

The deformation of large glass substrate in air-flotation system affects detection accuracy of inspection instrument. According to the gas lubrication theory, Timoshenko's thin film theory and the simulation figure of the pressure distribution of the air film flow field, the air load distribution model of the air film is established, and the deformation expression of the large liquid crystal glass substrate in air-flotation system is given. On this basis, a theoretical design method for designing nozzle pitch of orifice throttling air-flotation system was proposed. Combined with examples, the results of theories and simulation are compared. An experiment on the deformation of the glass substrate was carried out on an experimental prototype. The difference between the experimental results and the theoretical results does not exceed 10%.

Molecular genetic studies of gene identification for sarcopenia.

Sarcopenia, which is characterized by a progressive decrease of skeletal muscle mass and function with aging, is closely related to several common diseases (such as cardiovascular and airway diseases) and functional impairment/disability. Strong genetic determination has been reported for muscle mass and muscle strength, two most commonly recognized and studied risk phenotypes for sarcopenia, with heritability ranging from 30 to 85% for muscle strength and 45-90% for muscle mass. Sarcopenia has been the subject of increasing genetic research over the past decade. This review is designed to comprehensively summarize the most important and representative molecular genetic studies designed to identify genetic factors associated with sarcopenia. We have methodically reviewed whole-genome linkage studies in humans, quantitative trait loci mapping in animal models, candidate gene association studies, newly reported genome-wide association studies, DNA microarrays and microRNA studies of sarcopenia or related skeletal muscle phenotypes. The major results of each study are tabulated for easy comparison and reference. The findings of representative studies are discussed with respect to their influence on our present understanding of the genetics of sarcopenia. This is a comprehensive review of molecular genetic studies of gene identification for sarcopenia, and an overarching theme for this review is that the currently accumulating results are tentative and occasionally inconsistent and should be interpreted with caution pending further investigation. Consequently, this overview should enhance recognition of the need to validate/replicate the genetic variants underlying sarcopenia in large human cohorts and animal. We believe that further progress in understanding the genetic etiology of sarcopenia will provide valuable insights into important fundamental biological mechanisms underlying muscle physiology that will ultimately lead to improved ability to recognize individuals at risk for developing sarcopenia and our ability to treat this debilitating condition.

Tanshinone IIA induces growth inhibition and apoptosis in gastric cancer in vitro and in vivo.

As a phytochemical derived from the roots of Salvia miltiorrhiza Bunge, Tanshinone IIA has been reported to possess anti-inflammatory and antioxidant activity. Studies in breast, colon, prostate and lung cancer indicate that Tanshinone IIA may exhibit a promising antitumor activity. However, systemic studies of the cytotoxic effects of Tanshinone IIA on gastric cancer have not been described. The present study offers a comprehensive evaluation of the antitumor effects of Tanshinone IIA in gastric cancer cells in vitro and in a mouse xenograft model. Cell viability and apoptosis in vitro were evaluated through the MTT assay and flow cytometry analysis. The results indicate that Tanshinone IIA can induce gastric cancer cell growth inhibition and apoptosis in a time- and concentration-dependent manner. Furthermore, we investigated the mechanism of the apoptotic effects induced by Tanshinone IIA. We found that Tanshinone IIA can not only cause cell cycle arrest in the G2/M phase, but also trigger the intrinsic apoptotic signaling pathway. The results suggest that Tanshinone IIA may serve as an effective adjunctive reagent in the treatment of gastric cancer.

Copy number variation in CNP267 region may be associated with hip bone size.

Osteoporotic hip fracture (HF) is a serious global public health problem associated with high morbidity and mortality. Hip bone size (BS) has been identified as one of key measurable risk factors for HF, independent of bone mineral density (BMD). Hip BS is highly genetically determined, but genetic factors underlying BS variation are still poorly defined. Here, we performed an initial genome-wide copy number variation (CNV) association analysis for hip BS in 1,627 Chinese Han subjects using Affymetrix GeneChip Human Mapping SNP 6.0 Array and a follow-up replicate study in 2,286 unrelated US Caucasians sample. We found that a copy number polymorphism (CNP267) located at chromosome 2q12.2 was significantly associated with hip BS in both initial Chinese and replicate Caucasian samples with p values of 4.73E-03 and 5.66E-03, respectively. An important candidate gene, four and a half LIM domains 2 (FHL2), was detected at the downstream of CNP267, which plays important roles in bone metabolism by binding to several bone formation regulator, such as insulin-like growth factor-binding protein 5 (IGFBP-5) and androgen receptor (AR). Our findings suggest that CNP267 region may be associated with hip BS which might influence the FHL2 gene downstream.

Genome-wide association study identifies ALDH7A1 as a novel susceptibility gene for osteoporosis.

Osteoporosis is a major public health problem. It is mainly characterized by low bone mineral density (BMD) and/or low-trauma osteoporotic fractures (OF), both of which have strong genetic determination. The specific genes influencing these phenotypic traits, however, are largely unknown. Using the Affymetrix 500K array set, we performed a case-control genome-wide association study (GWAS) in 700 elderly Chinese Han subjects (350 with hip OF and 350 healthy matched controls). A follow-up replication study was conducted to validate our major GWAS findings in an independent Chinese sample containing 390 cases with hip OF and 516 controls. We found that a SNP, rs13182402 within the ALDH7A1 gene on chromosome 5q31, was strongly associated with OF with evidence combined GWAS and replication studies (P = 2.08x10(-9), odds ratio = 2.25). In order to explore the target risk factors and potential mechanism underlying hip OF risk, we further examined this candidate SNP's relevance to hip BMD both in Chinese and Caucasian populations involving 9,962 additional subjects. This SNP was confirmed as consistently associated with hip BMD even across ethnic boundaries, in both Chinese and Caucasians (combined P = 6.39x10(-6)), further attesting to its potential effect on osteoporosis. ALDH7A1 degrades and detoxifies acetaldehyde, which inhibits osteoblast proliferation and results in decreased bone formation. Our findings may provide new insights into the pathogenesis of osteoporosis.

The role of cellular oxidative stress in regulating glycolysis energy metabolism in hepatoma cells.

BACKGROUND: The Warburg effect has been found in a wide spectrum of human cancers, however the underlying mechanisms are still unclear. This study aims to explore the role of cellular oxidative stress in relation to glycolysis and the Warburg effect in hepatoma cells. METHODS: Various cell lines combining environmental hypoxia was used as an in vitro model to mimic tumor microenvironment in vivo. Superoxide dismutases (SOD) and xanthine oxidase (XO) gene transfection were used to produce various cellular redox levels. 2',7'-dichlorofluorescin (DCF) fluorescence and ESR spectrum were used to detect cellular reactive oxygen species (ROS). RESULTS: We found that endogenous or exogenous interference with the cellular oxidative stress can sensitively regulate glycolysis and the Warburg effect in hepatoma cells. Hepatoma cells displayed a high level of free radicals compared to immortalized normal hepatocyte cells. Increasing the level of ROS stress in hepatoma cells can directly upregulate HIF-1 and activate glycolysis without requirement of a hypoxic condition. This explains the mechanism whereby aerobic glycolysis, i.e. the Warburg effect arises. Either endogenously upregulating SOD or exogenously administration with antioxidant can, through downregulating ROS level, effectively regulate energy pathways in hepatoma cells and can inhibit the growth of tumor cells and xenograft tumors. CONCLUSION: This study suggests that the Warburg effect was related to an inherently high level of cellular ROS and HIF-1. Hepatoma cells adaptation to hypoxia for survival and rapid growth exploits oxidative stress ectopically activated glycolysis to compensate the energy supply. This specific mechanism in which tumor cells through cellular oxidative stress activate glycolysis to meet their energy metabolism requirement could be exploited to selectively kill tumor cells.

[The role of gap-junction in suspended animation for hemorrhagic shock with brain injury: experiment with rats].

OBJECTIVE: To investigate the role of gap-junction in suspended animation for hemorrhagic shock with brain injury. METHODS: Twenty-four SD rats were made into models of uncontrolled hemorrhagic shock and occlusion of bilateral common carotid arteries and randomly divided into 3 equal groups: traditional resuscitation group (Group I) undergoing traditional resuscitation, hypothermy resuscitation group (Group II) undergoing temperature lowering by ice-cap, and carbenoxolone resuscitation group (Group III) undergoing injection of carbenoxolone 50 microg/200 g. The process was divided into 3 periods: traumatic hemorrhagic period (30 min), pre-hospital treatment period (60 min), and in-hospital cardiopulmonary resuscitation period (60 min). The levels of left ventricular systolic pressure (LVSP), maximum change rate of left ventricular pressure rise and fall (+/- dp/dt(max)), mean arterial pressure (MAP), heart rate (HR), and respiratory rate (RR) were recorded at the beginning and the end of traumatic hemorrhagic period (T1 and T2), the end of pre-hospital treatment period (T3), and the end of in-hospital cardiopulmonary resuscitation period (T4). The survival time was recorded after in-hospital cardiopulmonary resuscitation period. Then the left brain was taken out and the hippocampal neurons apoptosis was observed by flow cytometry, chemiluminescence was used to detect the ATP, and IL-6 and tumor necrosis factor (TNF)-alpha were detected by enzyme linked immunosorbent assay (ELISA). RESULTS: The survival time of Groups II and III were (2.9 +/- 0.6) h and (2.6 +/- 1.0) h respectively, both significantly longer than that of Group I [(1.4 +/- 0.3) h, both P < 0.01]. The apoptotic rates of hippocampal neurons of Groups II and III were (72 +/- 6)% and (75 +/- 9)% respectively, both significantly lower than that of Group [83 +/- 5)% P < 0.05]. The ATP levels in hippocampus of Groups II and III were (2.0 +/- 0.3) and (1.9 +/- 0.4) pg/g respectively, both significantly higher than that of Group I [(1.4 +/- 0.5 pg/g, both P < 0.05). The TNF-alpha and IL-6 levels of Groups II were (1.7 +/- 0.3) pg/g and (19 +/- 3) pg/g respectively, both significantly lower than those of Group I [(2.2 +/- 0.6) and (24 +/- 3) pg/g respectively, both P < 0.05]. The IL-6 level of Group III was (26 +/- 4) pg/g, significantly higher than that of Group II (P < 0. 01). There were no significant difference between Groups II and III in the values of survival time, hippocampal neuron apoptosis, dissipation of ATP, and liberation of TNF-alpha (all P > 0.05), but there was significant difference in IL-6 (P < 0.01). CONCLUSION: Both hypothermy resuscitation and carbenoxolone resuscitation protect the brain of cerebral ischemia on hemorrhagic shock with brain injury, and suggest that Gap junctions play an important role in suspended animation to treat hemorrhagic shock with brain injury.

[Preparation of the spin trapping probe, N-tert-butyl-alpha-phenylnitrone, nanoparticle and its affinity to hepatoma cells].

This article describes the preparation of the N-tert-butyl-alpha-phenylnitrone (PBN) liposomes and their related characteristics. The PBN liposomes were prepared by film dispersion-supersonic method and the formula of liposomes was optimized by orthogonal uniform design. RP-HPLC was used to qualify the amount of PBN that entered into the hepatoma cells. Necrosis rate was also investigated by fluorescence activated cell sorter (FACS) after PBN liposomes transfection. Result showed that the mean particle size, entrapment efficiency, and polydispersity of the resulting PBN-liposome were 137.5 nm, 71.52% and 0.286, respectively. PBN liposomes can enter into the tumor cell stably and they have higher affinity to hepatoma cell compared with free PBN resulting in a higher necrosis rate after transfection. These results provide a potential method for early diagnosis and treatment of cancer using specific spin trapping probe targeting tumor cells.

Alpha-lipoic acid induces apoptosis in hepatoma cells via the PTEN/Akt pathway.

We report here that alpha-lipoic acid (alpha-LA), a naturally-occurring antioxidant, scavenges reactive oxygen species (ROS) followed by an increase in apoptosis of human hepatoma cells. Apoptosis induced by alpha-LA was dependent upon the activation of the caspase cascade and the mitochondrial death pathway. alpha-LA induced increases in caspase-9 and caspase-3 but had no significant effect on caspase-8 activity. Apoptosis induced by alpha-LA was found to be mediated through the tensin homologue deleted on chromosome 10 (PTEN)/Akt pathway. Prior to cell apoptosis, PTEN was activated and its downstream target Akt was inhibited. Our findings indicate that increasing ROS scavenging could be a therapeutic strategy to treat cancer.

N-[2-(2-Methoxyphenyl)benzylidene]-tert-butyl-amine N-oxide.

In the mol-ecule of the title compound, C(18)H(21)NO(2), the two benzene rings are oriented at a dihedral angle of 58.19 (3)°. Intra-molecular C-H⋯O hydrogen bonds result in the formation of one six- and one five-membered ring, which adopt twist and envelope conformations, respectively. In the crystal structure, C-H⋯O hydrogen bonds link the mol-ecules.

Formation of high-molecular-weight protein adducts by methyl docosahexaenoate peroxidation products.

In the present study, the formation of modified proteins by methyl docosahexaenoate (DHA) peroxidation products in the presence of a metal-catalyzed oxidation system was investigated. Metal-catalyzed oxidation of mixtures containing bovine serum albumin (BSA) and DHA led to formation of two high molecular weight derivatives of BSA. One had a mass of 71.5 kDa as determined by two-dimensional electrophoresis, matrix assisted laser desorption and ionization mass spectrometer (MALDI MS) analysis. The other was estimated to be 93 kDa by SDS-PAGE electrophoresis. The exposure of BSA to DHA also led to the generation of carbonyl groups. Oxygen radical scavengers could inhibit these modifications induced by DHA peroxidation. Furthermore, there was little difference of the peptides mass fingerprinting between the two kinds of modified high-molecular-weight proteins. These results suggest that oxygen radicals formed during lipid peroxidation are involved in the formation of protein derivatives. Our study may be important in the understanding the specific role of docosahexaenoic acid in the formation of modified proteins during aging and its related diseases.

Displacement of the beating heart induces an immediate and sustained increase in myocardial reactive oxygen species.

BACKGROUND: Heart manipulation and displacement are common maneuvers during beating heart surgery to expose coronary arteries for revascularization. Effects of heart displacement on free radical generation, reactive oxygen species (ROS) have not been previously described. METHODS AND RESULTS: Seven adult male dogs were anesthetized, a left lateral thoracotomy performed to expose the heart, and the coronary sinus cannulated for ROS sampling during different manipulation protocols: (1) heart in normal position; (2) 90 degree manual heart displacement; (3) Trendelenburg position while the heart displaced 90 degrees and (4) return heart to normal resting anatomical position and plus the operating table returned to horizontal. Heart displacement followed by anatomical re-positioning significantly increased the ROS signal as measured by EPR (50-fold compared with control values; p<0.01). CONCLUSION: Trendelenburg positioning and/or repositioning the heart during cardiac surgery may induce acute reperfusion injury and increase ROS.

Reduced cardiac output is associated with decreased mitochondrial efficiency in the non-ischemic ventricular wall of the acute myocardial-infarcted dog.

Cardiogenic shock is the leading cause of death among patients hospitalized with acute myocardial infarction (MI). Understanding the mechanisms for acute pump failure is therefore important. The aim of this study is to examine in an acute MI dog model whether mitochondrial bio-energetic function within non-ischemic wall regions are associated with pump failure. Anterior MI was produced in dogs via ligation of left anterior descending (LAD) coronary artery, that resulted in an infract size of about 30% of the left ventricular wall. Measurements of hemodynamic status, mitochondrial function, free radical production and mitochondrial uncoupling protein 3 (UCP3) expression were determined over 24 h period. Hemodynamic measurements revealed a > 50% reduction in cardiac output at 24 h post infarction when compared to baseline. Biopsy samples were obtained from the posterior non-ischemic wall during acute infarction. ADP/O ratios for isolated mitochondria from non-ischemic myocardium at 6 h and 24 h were decreased when compared to the ADP/O ratios within the same samples with and without palmitic acid (PA). GTP inhibition of (PA)-stimulated state 4 respiration in isolated mitochondria from the non-ischemic wall increased by 7% and 33% at 6 h and 24 h post-infarction respectively when compared to sham and pre-infarction samples. This would suggest that the mitochondria are uncoupled and this is supported by an associated increase in UCP3 expression observed on western blots from these same biopsy samples. Blood samples from the coronary sinus measured by electron paramagnetic resonance (EPR) methods showed an increase in reactive oxygen species (ROS) over baseline at 6 h and 24 h post-infarction. In conclusion, mitochondrial bio-energetic ADP/O ratios as a result of acute infarction are abnormal within the non-ischemic wall. Mitochondria appear to be energetically uncoupled and this is associated with declining pump function. Free radical production may be associated with the induction of uncoupling proteins in the mitochondria.

Reactive oxygen species stimulated human hepatoma cell proliferation via cross-talk between PI3-K/PKB and JNK signaling pathways.

Reactive oxygen species (ROS) are important for intracellular signaling mechanisms regulating many cellular processes. Manganese superoxide dismutase (MnSOD) may regulate cell growth by changing the level of intracellular ROS. In our study, we investigated the effect of ROS on 7721 human hepatoma cell proliferation. Treatment with H2O2 (1-10 microM) or transfection with antisense MnSOD cDNA constructs significantly increased the cell proliferation. Recently, the mitogen-activated protein kinases (MAPK) and the protein kinase B (PKB) were proposed to be involved in cell growth. Accordingly, we assessed the ability of ROS to activate MAPK and PKB. PKB and extracellular signal-regulated kinase (ERK) were both rapidly and transiently activated by 10 microM H2O2, but the activities of p38 MAPK and JNK were not changed. ROS-induced PKB activation was abrogated by the phosphatidylinositol 3-kinase (PI3-K) inhibitor LY294002, suggesting that PI3-K is an upstream mediator of PKB activation in 7721 cells. Transfection with sense PKB cDNA promoted c-fos and c-jun expression in 7721 cells, suggesting that ROS may regulate c-fos and c-jun expression via the PKB pathway. Furthermore we found that exogenous H2O2 could stimulate the proliferation of PKB-AS7721 cells transfected with antisense PKB cDNA, which was partly dependent on JNK activation, suggesting that H2O2 stimulated hepatoma cell proliferation via cross-talk between the PI3-K/PKB and the JNK signaling pathways. However, insulin could stimulate 7721 cell proliferation, which is independent of cross-talk between PI3-K/PKB and JNK pathways. In addition, H2O2 did not induce the cross-talk between the PI3-K/PKB and the JNK pathways in normal liver cells. Taken together, we found that ROS regulate hepatoma cell growth via specific signaling pathways (cross-talk between PI3-K/PKB and JNK pathway) which may provide a novel clue to elucidate the mechanism of hepatoma carcinogenesis.

[Influence of PKB on ROS regulation of proliferation in human 7721 hepatoma cells].

In the present study, the relationship between PKB signaling and reactive oxygen species (ROS) during the course of exogenous and endogenous ROS or antioxidants regulating human 7721 hepatoma cell proliferation was studied. To change endogenous ROS levels, 7721 cells were transfected with human manganese superoxide dismutase (MnSOD) construct containing sense or antisense MnSOD cDNA. Low level of exogenous ROS H2O2(1-10 mumol/L) significantly stimulated PKB activity and c-fos/c-jun expression and cell growth, which could be abolished by antioxidant danshensu (40 mg/L). It was observed that overexpression of MnSOD inhibited 7721 cell growth by inhibiting PKB activity and c-fos/c-jun expression; the PKB activity and c-fos/c-jun expression, however, were stimulated by down-regulated MnSOD expression. In addition, PKB-7721 cells (transfected with sense PKB cDNA) promoted c-fos/c-jun expression by stimulating PKB activity. These results suggest that the redox state stimulated hepatoma cell growth through PKB pathway, which modulates AP-1 expression.

Inhibition of Proliferation and Expression of N-ras in Hepatoma Cells by Antioxidation Treatment.

Kunming mice inoculated with hepatoma cell (H22) suspension subcutaneously at their right axilla were administered orally with antioxidants such as vitamine E, beta-carotene, glutamine, kappa-selenocarrageenan and polysaccharide-peptide of coriolus (PSP) solution. It was found that the inoculated hepatoma growth was suppressed to various extents. The two kinds of polysaccharide antioxidants improved non-specific immunity, enhanced the nitrogen monoxide (NO) content in plasma and strengthened the inhibition of hepatoma. Above antioxidants added in the culture of 7721 human hepatoma cells inhibited the cell proliferation and inducedits apoptosis. Meanwhile, the activity of glutathione peroxidase (GSH-Px) in the plasma of mice increased and the content of malondialdehyde (MDA) decreased. H(2)O(2) in low concentration improved the cancer cell proliferation and inhanced the expression of Mn-SOD c-fos and c-jun, but led to cells apoptosis or necrosis in high concentration. The mechanism of antioxidants inhibiting tumor growth and improving cancer cells apoptosis might be that, on the one hand, the antioxidants blocked the free radicals signal transduction on cancer cells proliferation, and on the other hand, they improved the release of NO through enhancing the non-specific immunity, so inhibiting the cancer cells proliferation directly.